A biosensor is a device, probe, or electrode which, when in contact with an appropriate sample, produces an electrical signal in the presence of the desired analyte. The biosensor is normally constructed by immobilizing a biologically sensitive material in intimate contact with a suitable transducing system to convert the concentration of the analyte into a quantifiable signal.
In spite of several advantages, the currently available biosensors all suffer from many problems awaiting solution: chemical interference, environmental effects, long-term stability, signal-to-noise ratio and design of the sensor's packaging system.
The following trends are recognizable in the development of biosensors:
a) Miniaturization
b) Determination of several reagents with an array sensor system combining more than one sensing element
c) Development of mass producible disposable sensors
d) In vivo analysis using implantable biochips    e) Processing of the output from an arrayed sensor system of with an artificial intelligence system
Meanwhile, the development of a rapid, simple, separation-free method for the detection of proteins has been a long-standing goal. Chromogenic and fluorogenic galactoside-dextran substrates have been used to devise homogeneous enzyme immunoassays (EIAs) for C-reactive protein, ferritin, and immunoglobulins (Gibbons et al., “Homogeneous Enzyme Immunoassay for Proteins Employing β-Galactosidase,” Analytical Biochemistry 102/167-170, 1980; and Armenta et al., “Improved Sensitivity in Homogeneous Enzyme Immunoassays Using a Fluorogenic Macromolecular Substrate: An Assay for Serum Ferritin,” Analytical Biochemistry 146/211-219, 1985). However, the low degree of modulating enzyme activity in this homogeneous protocol has rendered the method impractical for real world applications.
Also, a separation-free dual solid-phase EIA for macromolecules, which relies on the partitioning of an enzyme conjugate (biotin-glucose-6-phosphate dehydrogenase-antibody) between two solid phases of polystyrene latex-bound avidin and polystyrene latex-bound analyte, has been reported (Schray et al., “Separation-Free Dual Solid Phase Enzyme Immunoassay for Macromolecules,” Analytical Chemistry, 60/353-56 1988). However, this assay scheme requires 24 hours for enzymatic generation of a detectable product.
It has long been recognized that coupling electrochemical detection with EIAs would be advantageous. Electrodes are insensitive to the color or turbidity of a test sample and can therefore be used to develop methods directly applicable to whole blood samples. However, most of the many reports regarding the use of electrochemical detection to devise EIAs or “immunosensors” have relied on using such sensors as solid phases in heterogeneous assay arrangements in which antibodies are immobilized at the surface of a given electrode. After incubation of a sample with other reagents, the surface of the electrodes has to be washed before adding the substrate needed to measure bound enzyme activity.
As a specific example, in U.S. Pat. No. 5,063,081 issued Nov. 5, 1991, Cozzette et al. disclose a ligand/ligand receptor-based biosensor for detecting a particular analyte species, such as an antigen. Here, a base sensor, comprising a catalytic indicator electrode using a noble late transition metal such as iridium, gold, platinum, or silver, is surrounded by a combined reference and counter electrode made of, for example, silver and silver chloride (columns 25-26). An antibody is immobilized on the base sensor. The resulting biosensor is then brought into contact with a mixture comprising the sample and a second analyte-specific antibody, which is labeled (columns 45-46). A permselective silane layer may also be used as a screen against interfering species. However, unbound materials and interfering electroactive species are preferably removed from the sensor by using either a wash solution or by using the solution containing the enzyme substrate as a wash (columns 47-49).
As another specific example, in U.S. Pat. No. 5,830,680 issued Nov. 3, 1998, Meyerhoff et al. disclose an enzyme sandwich immunoassay cassette for detecting an analytical signal over any background signal originating from a bulk solution in contact with the cassette. Here, the cassette comprises a microporous membrane support having coated on one side thereof a conductive metal layer and at least a first capture antibody layer immobilized over the conductive metal layer in at least a first spatially distinct area of the microporous membrane support. Referring to FIG. 1, which is a diagram of a diffusion cell arrangement, it can be seen that the cassette requires an additional auxiliary electrode and/or a reference electrode, such that miniaturization and point-of-care testing are not achieved.
Therefore, despite all of the past and current research activity in this area, a new biosensor that avoids the above-described disadvantages has long been desired.